1. Field of the Invention
The present invention relates to a mutant strain of Bacillus subtilis and its application in preparing acetoin, especially relates to a mutant strain of Bacillus subtilis CGMCC No. 1869 and its application in producing acetoin of high purity.
2. Description of the Related Art
Acetoin (acetylmethylcarbinol) widely exists in corn, grape, apple, banana, cheese, meat and many other foods. Acetoin contributes the special butter flavor to all these foods. And beer flavor is also related to acetoin.
Acetoin is a broadly used as one of the food flavors in the world, and it is also an important chemical compound and a medicine intermediate. Acetoin can be used to synthesize diacetyl and 2,3-butanediol. As a chiral compound, it can be used to synthesize some rare medicines and medical intermediates, such as 4-chloro-4,5-dimethyl-1,3-dioxolan-2-one(CDMDO), an important intermediate mainly used for modifying penicillin, ampicillin and other antibiotics to increase their activities and decrease their medicine side effects. In addition, acetoin can also be used to synthesize lenampicillium hydrochloride (X. H. Xiao, 2004), a precursor of half synthetic penicillin and ampicillin. Lenampicillium hydrochloride is stable in intestines and stomach, of better absorption, less bad effects and low toxicity.
Acetoin can be synthesized chemically by partial hydrogenated reduction of diacetyl (2,3-butanedione) or selective oxidization by 2,3-butanediol with the transformation efficiency of 50-80%. Acetoin can also be prepared by electronoxidizing 2,3-butanediol with better selectivity and high yield (A. Hilmi, et al, 1997). Besides, acetoin can be produced by enzymatic conversion. For examples, an U.S. Pat. No. 5,164,314 disclosed a process for the enzymatic preparation of acetoin, which employed a diacetyl reductase from a yeast or Lactobacillus strain (Hummel in 1992); Susan Budavari reported that sorbose bacterium could transform 2,3-butanediol to acetoin with high yield without by-products; however, it is very complicated for addressing those specific enzymes involved in this process.
Acetoin is an important metabolite in some microorganisms, so that it also can be produced by microbial fermentation by consuming glucose. However, most reports relevant to acetoin fermentation were about the metabolic pathways and only a few of reports were related to acetoin production as a by-product of butanedione or 2,3-butanediol. In U.S. Pat. No. 5,075,226, a method was disclosed for production of diacetyl and acetoin using lactic acid bacterium by fermentation. Hespell reported that Bacillus polymyxa was able to utilize xylan to produce butanedione and acetoin and the maximal total yield reached to 11.3 g/L. Braneni & Keenan found diacetyl and acetion can be produced by fermentation using lactic acid bacteria, which also produced 2,3-butanediol simultaneously. Teixeira reported that Hanseniaspora guilliermondii could produce acetoin with a yield as high as to 0.36 g/L. In present invention the inventors provide a Bacillus subtilis mutant strain which can produce acetoin with advantages far beyond all above.